Previously, we reported that levels of chymase activity and its mRNA in cardiac tissues were significantly increased along with progression of cardiac fibrosis in cardiomyopathic hamsters, but the involvement of chymase in the progression of fibrosis has been unclear. In cultured human fibroblasts, the concentration of transforming growth factor-␤ in the supernatant of medium was significantly increased after injection of human chymase. Furthermore, human chymase dose dependently increased cell proliferation, and this chymase-dependent proliferation was completely suppressed by a chymase inhibitor, Suc-Val-Pro-Phe p (OPh) 2 (10 M) or an anti-transforming growth factor-␤ antibody (100 g/ml). In this study, we used Bio14.6 and F1B hamsters as cardiomyopathic and control hamsters, respectively. Cardiomyopathic hamsters were orally administered a novel chymase inhibitor, 4-[1-{[bis-(4-methylphenyl)-methyl]-carbamoyl}-3-(2-ethoxy-benzyl)-4-oxo-azetidine-2-yloxy]-benzoic acid (BCEAB; 100 mg/kg per day), or placebo from 5-to 45-week-old. In the placebo-treated group, the cardiac chymase activity in cardiomyopathic hamsters 45 weeks old was significantly increased compared with that in control hamsters. BCEAB significantly reduced the cardiac chymase activity. The indexes (ϩdP/dt and -dP/dt) of cardiac function were significantly improved by treatment with BCEAB. The mRNA levels of collagen I and collagen III in the placebotreated hamsters were significantly reduced to 69.6 and 76.5% by treatment with BCEAB, respectively. The fibrotic area in cardiac tissues in the BCEAB-treated hamsters was significantly suppressed to 50.7% compared with that in the placebo-treated treated hamsters. Therefore, the activation of transforming growth factor-␤ by chymase may play an important role in the progression of cardiac fibrosis and cardiac dysfunction in cardiomyopathy.

A local segmental mobility was determined by electron spin resonance (ESR) spin-label method for a series of polystyrene (PS) with various molecular weights. Each PS specimen was selectively spin-labeled with stable nitroxide radicals at a chain end or inside sites. Molecular motion at the inside of the chain was compared with that at the chain end from the temperature dependence of ESR spectra of the nitroxide radicals. The transition temperature of molecular motion, T 5.0mT, at which the extreme separation width due to 14 N anisotropic hyperfine splitting is 5.0 mT, increased with an increase in molecular weight. The WLF equation confirmed that the T5.0mT correlated with a glass transition temperature, Tg, of PS. The T5.0mT for the spin-labeled PS at the chain end was ca. 5 K lower than that for the spin-labeled PS at the inside sites due to the enrichment of the specific free volume around the chain end. The transition temperature, T5.0mT, for both labeled PS depended on the molecular weight in accordance with the Unberreiter-Kanig equation for a glass transition. The T5.0mT for the spin-labeled PS at the chain end had a strong dependence on the molecular weight as compared with that at the inside sites because the molecular motion of the chain end was accelerated by an encounter of more than two chain ends. From the molecular weight dependence, we determined the short correlation time for segmental motion of the chain end, ca. 40 s, and the segment size undergoing the segmental motion at the T g. The obtained segment size agreed well with the general segment size reported by others, 5-10 monomeric unit size.

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